Instituto de Investigaciones Bioquímicas de La Plata (INIBIOLP), CCT- La Plata, CONICET, Facultad de Ciencias Médicas, Universidad Nacional de La Plata. 60 y 120, 1900 La Plata, Argentina.
Consejo Nacional de Investigaciones Científicas y Técnicas, Instituto de Química y Fisico-Química Biológicas (IQUIFIB) Prof. Alejandro C. Paladini, Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, Junín 956, Buenos Aires, Argentina.
Biochem J. 2019 Nov 29;476(22):3455-3473. doi: 10.1042/BCJ20190546.
Alpha hemolysin (HlyA) is the major virulence factor of uropathogenic Escherichia coli (UPEC) strains. Once in circulation, a low concentration of the toxin induces an increase in intracellular calcium that activates calpains - which proteolyse cytoskeleton proteins - and also favours the exposure of phosphatidylserine (PS) in the outer leaflet of erythrocyte membranes. All these events are considered part of eryptosis, as well as the delivery of microvesicles (MVs). Within this context, we studied the delivery of MVs by erythrocytes treated with sublytic concentrations of HlyA and demonstrated that HlyA-treated erythrocytes secrete MVs of diameter ∼200 nm containing HlyA and PS by a mechanism involving an increment of intracellular calcium concentration and purinergic receptor activation. Despite the presence of toxin in their membrane, HlyA-MVs are not hemolytically active and do not induce ATP release in untreated erythrocytes, thus suggesting that the delivery of HlyA-MVs might act as a protective mechanism on the part of erythrocytes that removes the toxin from the membrane to prevent the spread of infection. Although erythrocytes have been found to eliminate denatured hemoglobin and several membrane proteins by shedding MVs, the present work has revealed for the first time that an exogenous protein, such as a toxin, is eliminated by this process. This finding sheds light on the mechanism of action of the toxin and serves to further elucidate the consequences of UPEC infection in patients exhibiting HlyA-related diseases.
α-溶血素（HlyA）是尿路致病性大肠杆菌（UPEC）菌株的主要毒力因子。一旦进入循环，毒素的低浓度会引起细胞内钙离子增加，激活钙蛋白酶 - 裂解细胞骨架蛋白 - 并有利于红细胞膜外层磷脂酰丝氨酸（PS）的暴露。所有这些事件都被认为是细胞发生凋亡的一部分，以及微泡（MVs）的释放。在这种情况下，我们研究了用亚致死浓度的 HlyA 处理的红细胞释放 MVs 的情况，并证明 HlyA 处理的红细胞通过增加细胞内钙离子浓度和嘌呤能受体激活来分泌直径约为 200nm 的含有 HlyA 和 PS 的 MVs。尽管其膜中存在毒素，但 HlyA-MVs 没有溶血活性，也不会在未处理的红细胞中诱导 ATP 释放，因此表明 HlyA-MVs 的释放可能是红细胞的一种保护机制，将毒素从膜中去除，以防止感染的扩散。尽管已经发现红细胞通过释放 MVs 来消除变性血红蛋白和几种膜蛋白，但本工作首次揭示了一种外源性蛋白质，如毒素，也可以通过这种过程被消除。这一发现揭示了毒素的作用机制，并有助于进一步阐明在表现出与 HlyA 相关疾病的患者中 UPEC 感染的后果。
